Figure S2.
Structure and function of ER exit sites in BFA/Noc treated cells. (A) Relative localization of Sec24C-mCherry (red) and VSVG (green) within ERES of BFA/Noc-treated COS7 cells. Top, left to right: (1) Confocal microscope. (2) Immunofluorescence analysis of endogenous Sec24C (red) in a cell expressing VSVG-YFP (green). (3) Cells under Golgicide A/Colchicin treatment. (4) Deconvolution using DeltaVision microscope. Bottom, left to right: (1) Multifocal structured illumination microscopy (M-SIM). (2) Relative localization of Sec24C-mCherry (red) and ERGIC53-YFP, GalT-YFP, or CFTR-GFP (green) within a representative ERES of COS7 cells under BFA/Noc treatment. Scale bars = 1 µm. (B) Cargo and membrane accumulation in a single dilated exit site. A COS7 cell transfected with cargo protein VSVG-YFP and treated with BFA/Noc was monitored by a spinning disk confocal microscope captured at 5 frames/s. Representative time-lapse images of a single dilated exit site marked by the white square frame, are shown in the middle panel. Bottom: A kymogram generated from the line marked by the yellow line. Scale bar = 1 µm. (C) Analysis of COPII-mediated cargo sorting dynamics in ERESs of BFA/Noc-treated living COS7 cells. A cell coexpressing VSVG-YFP (green) and Sec24C-mCherry (red) was imaged using a spinning disk confocal microscope at 0.2-s intervals at the permissive temperature 32°C. The fluorescence intensity of VSVG-YFP (green line) and Sec24C-mCherry (red line) in three representative ERESs I through III (in white frames) are plotted, and representative images are shown on the right. Black arrowheads in the graph show time points of images on the right. Scale bar = 5 µm. (D) The mutant cargo protein CFTRΔ508-GFP (green, bottom) is excluded from ERES in BFA/Noc-treated COS7 cells. The cells are coexpressing Sec24C-mCherry (red) with either WT CFTR-GFP (green, top) or CFTRΔ508-GFP (green, bottom). Insets are enlarged fivefold. Left scale bars = 10 µm. Right scale bars = 1 µm.

Structure and function of ER exit sites in BFA/Noc treated cells. (A) Relative localization of Sec24C-mCherry (red) and VSVG (green) within ERES of BFA/Noc-treated COS7 cells. Top, left to right: (1) Confocal microscope. (2) Immunofluorescence analysis of endogenous Sec24C (red) in a cell expressing VSVG-YFP (green). (3) Cells under Golgicide A/Colchicin treatment. (4) Deconvolution using DeltaVision microscope. Bottom, left to right: (1) Multifocal structured illumination microscopy (M-SIM). (2) Relative localization of Sec24C-mCherry (red) and ERGIC53-YFP, GalT-YFP, or CFTR-GFP (green) within a representative ERES of COS7 cells under BFA/Noc treatment. Scale bars = 1 µm. (B) Cargo and membrane accumulation in a single dilated exit site. A COS7 cell transfected with cargo protein VSVG-YFP and treated with BFA/Noc was monitored by a spinning disk confocal microscope captured at 5 frames/s. Representative time-lapse images of a single dilated exit site marked by the white square frame, are shown in the middle panel. Bottom: A kymogram generated from the line marked by the yellow line. Scale bar = 1 µm. (C) Analysis of COPII-mediated cargo sorting dynamics in ERESs of BFA/Noc-treated living COS7 cells. A cell coexpressing VSVG-YFP (green) and Sec24C-mCherry (red) was imaged using a spinning disk confocal microscope at 0.2-s intervals at the permissive temperature 32°C. The fluorescence intensity of VSVG-YFP (green line) and Sec24C-mCherry (red line) in three representative ERESs I through III (in white frames) are plotted, and representative images are shown on the right. Black arrowheads in the graph show time points of images on the right. Scale bar = 5 µm. (D) The mutant cargo protein CFTRΔ508-GFP (green, bottom) is excluded from ERES in BFA/Noc-treated COS7 cells. The cells are coexpressing Sec24C-mCherry (red) with either WT CFTR-GFP (green, top) or CFTRΔ508-GFP (green, bottom). Insets are enlarged fivefold. Left scale bars = 10 µm. Right scale bars = 1 µm.

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